JP4534087B2 - Endotoxin adsorption remover and production method - Google Patents

Description

  The present invention relates to an adsorption / removal agent effective for removing a substance that induces fever, that is, a pyrogen (pyrogen) when mixed in a living body.

  In particular, the present invention relates to an endotoxin adsorption / removal agent suitable for adsorbing and removing endotoxins mixed in liquids such as water, injection solutions, infusion solutions, dialysis solutions, and blood.

  Moreover, this invention relates also to the manufacturing method of the said endotoxin adsorption removal agent.

  Endotoxin is one of the outer membrane constituents corresponding to the epidermis of the cell wall of Gram-negative bacteria, and is released into the solution when it is lysed, mechanically destroyed, or when the bacteria divide.

  When such endotoxins are mixed into animal blood, they cause toxic effects such as fever, inflammation, and death from shocks. Therefore, various measures have been taken to remove these endotoxins.

  For example, conventional methods for removing or deactivating endotoxins include treatment with heat, acid, alkali, etc., but these methods are time-consuming and are not methods for removing only endotoxins, and therefore, coexisting active ingredients, etc. Are also likely to be removed or deactivated.

  Therefore, in recent years, the development of adsorbents taking advantage of the removal characteristics at the molecular weight level using an ultrafiltration membrane or the like, and the adsorption characteristics of the material has been active. For example, porous silica gel having an amino group and a hydrophobic group obtained by reacting a porous silica gel with a silane coupling agent having an amino group and a hydrophobic group (see, for example, Patent Document 1), silicon oil, silicon on the inorganic surface Endotoxin adsorption / removal agent using a polymer or paraffin-attached composite material (see, for example, Patent Document 2 or 3), water-insoluble calcium salts such as calcium carbonate (calcium carbonate, calcium phosphate, hydroxyapatite) ( For example, refer to Patent Document 4), and endotoxin adsorption / removal agents using oxide ceramics such as titania and zirconia as an adsorbent (for example, refer to Patent Document 5).

However, these adsorbent removers require a large amount of sample to adsorb endotoxin, and in order to process high-concentration endotoxin solution and a large amount of solution, a large number of adsorbent removers or large-sized devices are required. There is a problem such as having to use.
JP-A-4-256434 (Claims) JP-A-7-136506 (Claims) JP-A-7-136507 (Claims) JP-A-7-265691 (Claims) JP-A-10-235188 (Claims)

  An object of the present invention is to solve the above-mentioned problems and to provide an adsorption / removal agent having excellent adsorption / removability against endotoxin and a method for producing such an adsorption / removal agent.

  As a result of intensive studies in view of the above-mentioned problems of the prior art, the present inventor found that an inorganic compound containing a specific aluminum silicate as an active ingredient removes endotoxin extremely effectively, and at low cost and endotoxin removal efficiency. An excellent adsorption remover has been completed. The present inventor has also found that by using colloidal silica as a silicon source, aluminum silicate capable of removing endotoxin extremely effectively can be produced. That is, this invention relates to the following endotoxin adsorption removal agent and its manufacturing method.

Item 1. An endotoxin adsorption / removal agent characterized by containing aluminum silicate as an active ingredient and having an electrokinetic potential (zeta potential) when the aluminum silicate is dispersed in an aqueous solvent as a positive charge .

Item 2 . Item 2. The method for producing an endotoxin adsorption / removal agent according to Item 1 , comprising a step of obtaining an aluminum silicate coprecipitate using colloidal silk as an aluminum salt and a silicon source.

  The endotoxin adsorption / removal agent of the present invention is excellent in endotoxin adsorption performance, and can effectively reduce the toxic effect of endotoxin.

  Such an endotoxin adsorption / removal agent of the present invention can exhibit an endotoxin adsorption / removal action even in a small amount, in addition to its active component being inexpensive. Furthermore, endotoxin can be adsorbed and removed by a simple operation without using a large apparatus or the like. Therefore, by using the adsorption removing agent of the present invention, endotoxin-free water, injection solution, infusion solution, dialysis solution, blood and the like can be efficiently provided at low cost.

  Furthermore, according to the production method of the present invention, an endotoxin adsorption / removal agent that is excellent in endotoxin adsorption performance and can efficiently reduce the toxic effect of endotoxin can be produced.

  First, the aluminum silicate which the endotoxin adsorption removal agent of this invention contains as an active ingredient is demonstrated in detail.

The aluminum silicate which is an active ingredient of the endotoxin adsorption removing agent of the present invention is an aluminum silicate generally represented by a composition formula of xAl ₂ O ₃ · SiO ₂ · yH ₂ O. This aluminum silicate is a compound of silica / alumina and has an amorphous state that does not show a clear crystal structure. In the aluminum silicate used in the present invention, the ratio of aluminum (Al) to silicon (Si) (hereinafter also referred to as “Al / Si ratio”) includes both silicon and aluminum, and adsorbs endotoxin. There is no particular limitation as long as it can be removed. Specifically, the Al / Si ratio can be used from a value exceeding 0 to about 50, but in general, the Al / Si ratio is from about 0.2 to around 25. Can be used. The Al / Si ratio is obtained from the weight of silicon dioxide and the weight of aluminum oxide obtained by the silicon dioxide (SiO ₂ ) quantitative method and aluminum oxide (Al ₂ O ₃ ) quantitative method described in Example 1 described later. The molar ratio between Al and Si.

  The aluminum silicate used in the present invention has a positive charge on the electrokinetic potential (zeta potential) (hereinafter also simply referred to as “zeta potential”) when it is dispersed in purified water. This is preferable from the viewpoint of enhancing the adsorption removal performance. When the aluminum silicate used in the present invention has a positive zeta potential, the mechanism of action for improving the adsorption removal performance with endotoxin is not necessarily clear. However, considering that the zeta potential is near the particle surface and the structure of endotoxin (consisting of an O antigen site, a core polysaccharide site, and a lipid A site, the lipid A site contains a long chain fatty acid). The attractive force between the positive charge near the particle surface and the negative ion of the fatty acid residue of endotoxin is considered to be one of the mechanisms of action. Based on this mechanism of action, it is predicted that a larger positive value of the zeta potential of aluminum silicate is preferable for improving the adsorption performance. In addition, it is predicted that the aluminum silicate used in the present invention can also adsorb substances derived from bacteria having a COOH group (tycoic acid).

  In the present invention, the zeta potential is determined when the zeta potential is dispersed in purified water because the zeta potential varies depending on the ions in the dispersion medium (such as the type and amount of coexisting ions including hydrogen ions). is there. Furthermore, the zeta potential can be changed by the water temperature, viscosity, and the like. Therefore, in the present invention, the zeta potential is measured under the conditions described in Example 1 described later.

  As described above, the zeta potential of a particle varies depending on the environment in which it is dispersed (such as the type and amount of coexisting ions including hydrogen ions). Describing the relationship between aluminum silicate that can be used in the present invention and pH, the zeta potential has a large absolute value of positive charge at the pH value on the acidic side, and becomes absolute as the pH value changes from the acidic side to the alkaline side. As the value decreases and goes through 0, and then the pH value further shifts to the alkaline side, the zeta potential becomes a negative charge and its absolute value increases. The endotoxin adsorption / removal agent of the present invention is used not only when it is dispersed in purified water, but also in the environment in which it is used (for example, injection solution, infusion solution, dialysis solution, and water for preparing them). It is preferable to have an electric charge in order to exhibit endotoxin adsorption removal performance.

  Although the aluminum silicate used by this invention can be manufactured with the following method, it is not limited to what is manufactured with this manufacturing method.

  As an example of the method for producing aluminum silicate used in the present invention, there can be mentioned a method in which a substance serving as an aluminum source and a substance serving as a silicate source are reacted successively or simultaneously at room temperature. As the aluminum source, aluminum salts such as aluminum sulfate and aluminum chloride and aluminates can be used. On the other hand, as the silicic acid source, colloidal silica, silica sol, wet silica gel, sodium silicate liquid (also called water glass), or the like can be used.

  Among the above-mentioned silicate sources, in particular, by using colloidal silica, aluminum silicate having higher endotoxin adsorption removal performance can be produced. The reason is not necessarily clear, but by adding a solution-like aluminum salt to colloidal silica, the composition of aluminum and silica becomes uniform, and the aluminum silicate synthesized in such a reaction system has its This is probably because a uniform composition in which aluminum and silica are not present alone in the structure is obtained. On the other hand, in a compound in which aluminum and silica exist alone, since the zeta potential of silica or the like is negative, there is a possibility that the zeta potential is totally negative. Presumed to decline.

  As for pH at the time of reaction with an aluminum source and a silicic acid source, 3-7 are preferable, and especially 4-6 vicinity is preferable. This is because elution of Al and Si is large in extreme acidity or alkalinity. For pH adjustment, when it is consumed as an acid or alkali such as sodium carbonate, sodium hydroxide, hydrochloric acid, carbon dioxide, etc., it is difficult for residual ions to react with silicic acid or aluminum, and it can be easily removed by washing with water Is good.

  Through the steps described above, the aluminum silicate used in the present invention is obtained as a precipitate. This precipitate is subjected to solid-liquid separation by filtration or the like, and washed (for example, washed with water) and dried as necessary to obtain aluminum silicate in a dry state. Although drying can be performed by a conventional method, for example, 200 ° C. or less, preferably 40 to 100 ° C. is preferable.

  The adsorption / removal agent of the present invention is used by adding aluminum silicate as an active ingredient as it is in a powder state to a liquid that can contain endotoxin (for example, water, injection solution, infusion solution, dialysis solution, blood, etc.). (So-called batch method). This batch method is particularly suitable when processing high concentrations of endotoxin. In addition, when it is necessary to physically remove the adsorption / removal agent of the present invention that has adsorbed endotoxin from the liquid for legal reasons or for other reasons, it can be separated and removed by filtration or the like.

  On the other hand, when endotoxin is continuously removed in a large amount, a column method is appropriate, and an adsorption removing agent is filled in the column, and a target solution or the like may be passed through the column.

  Moreover, the adsorption removal agent of this invention can also be used as an adsorption removal agent composition like a granulated material using excipient | fillers, such as a cellulose type and alginic acid type, as needed.

  Furthermore, the adsorptive removal agent of the present invention can be used by dispersing it in a solvent such as water or alcohol as needed and placing it on a nonwoven fabric or paper to carry the agent.

  Examples of the present invention will be described below, but the present invention is not limited to these examples. In the examples described below, “%” is% by weight unless otherwise specified.

Example 1
400 g of aluminum sulfate solution (about 8% as Al ₂ O ₃ ) and colloidal silica (about 20% as SiO ₂ , Snowtex 20 (product name) manufactured by Nissan Chemical Industries, Ltd., the same in the following examples) 37 g Were mixed with stirring to prepare a mixed solution. The total amount of the above mixture and 154 g of sodium hydroxide (about 48%) were added dropwise to 200 mL of water so that the pH was about 5.

  The produced precipitate was filtered, sufficiently washed with water, taken out, and dried by a conventional method to obtain a dried product.

The aluminum oxide content (as Al ₂ O ₃ ) and silicon dioxide content (as SiO ₂ ) in the obtained dried product were analyzed by the following procedures, and values were obtained. In addition, the zeta potential in the supernatant of a suspension (0.1%) in which 0.1 g of this product was suspended in 100 mL of purified water was measured using a zeta potential measuring device (manufactured by Nano-zs MALVERN). Note that Henry's formula was used to calculate the zeta potential. As a result, it was 39.42% as Al ₂ O _{3 and} 7.07% as SiO ₂ , and the molar ratio (Al / Si) was 6.55. The zeta potential was 13.41 mV.

<Quantitative determination of silicon dioxide (SiO ₂ )>
Weigh exactly 1.0 g of this product, add 20 mL of diluted hydrochloric acid, and evaporate to dryness on a water bath. The residue is moistened with hydrochloric acid and evaporated to dryness again. Add 5 mL of dilute hydrochloric acid to the residue and stir. Add 20 mL of hot water, stir and filter. The precipitate on the filter paper is washed thoroughly with hot water, transferred to a crucible of known weight together with the filter paper, converted to heat ash, ignited at 800 ° C. for 1 hour, allowed to cool, and then weighed to determine the amount of SiO ₂ .

  In addition, the filtrate and the washing solution are combined, and water is added to make exactly 250 mL as the sample solution for aluminum oxide determination.

<Aluminum oxide (Al ₂ O ₃ ) determination method>
Accurately measure 5 mL of the sample solution, add exactly 30 mL of 0.01M ethylenediaminetetraacetic acid disodium solution, add 20 mL of pH4.8 acetic acid / ammonium acetate buffer, boil for 5 minutes, cool, and add 50 mL of ethanol. Titrate with 0.01M zinc acetate solution (indicator: dithizone test solution 2mL). However, the end point of titration is when the color of the liquid changes from light green to light red. A blank test is performed in the same manner.

0.01M ethylenediaminetetraacetic acid disodium solution 1mL = 0.5098mg Al ₂ O ₃
(Example 2)
400 mL of an aluminum sulfate solution (about 8% as Al ₂ O ₃ ) and 12.5 g of colloidal silica (about 20% as SiO ₂ ) were stirred and mixed to prepare a mixed solution. The total amount of the above mixture and 164 g of sodium hydroxide (about 48%) were simultaneously added dropwise to 200 mL of water so that the pH was about 5.

  The produced precipitate was filtered, sufficiently washed with water, taken out, and dried by a conventional method to obtain a dried product.

The aluminum oxide content (as Al ₂ O ₃ ), silicon dioxide content (as SiO ₂ ), and zeta potential in the obtained dried product were analyzed in the same procedure as in Example 1 to determine values. As a result, Al ₂ O ₃ was 41.19%, SiO ₂ was 2.31%, and the molar ratio (Al / Si) was 21.79. The zeta potential was 14.81 mV.

(Practice interest 3)
200 mL of aluminum sulfate liquid (about 8% as Al ₂ O ₃ ) and 500 g of colloidal silica (about 20% as SiO ₂ ) were mixed with stirring to prepare a mixed solution. The total amount of the above mixture and 80 g of sodium hydroxide (about 48%) were added dropwise to 200 mL of water so that the pH was about 5.

  The produced precipitate was filtered, sufficiently washed with water, taken out, and dried by a conventional method to obtain a dried product.

The aluminum oxide content (as Al ₂ O ₃ ), silicon dioxide content (as SiO ₂ ), and zeta potential in the obtained dried product were analyzed in the same procedure as in Example 1 to determine values. As a result, Al ₂ O ₃ was 13.16%, SiO ₂ was 66.83%, and the molar ratio (Al / Si) = 0.23. The zeta potential was 12.00 mV.

Example 4
423 mL of aluminum chloride liquid (about 10% as Al ₂ O ₃ ) and 37 g of colloidal silica (about 20% as SiO ₂ ) were stirred and mixed to prepare a mixed solution. The total amount of the above mixture and 182 g of sodium hydroxide (about 48%) were simultaneously added dropwise to 200 mL of water so that the pH was about 5.

  The produced precipitate was filtered, sufficiently washed with water, taken out, and dried by a conventional method to obtain a dried product.

The aluminum oxide content (as Al ₂ O ₃ ), silicon dioxide content (as SiO ₂ ), and zeta potential in the obtained dried product were analyzed in the same procedure as in Example 1 to determine values. As a result, the Al ₂ O ₃ was 47.70%, the SiO ₂ was 8.13%, and the molar ratio (Al / Si) was 6.93. The zeta potential was 47.52 mV.

(Test Example 1)
Examples 1 to 4 and comparative example: Calcium silicate (as in Example 1, aluminum oxide content (as Al ₂ O ₃ ), silicon dioxide content (as SiO ₂ ) and zeta potential , Al ₂ O ₃ as a 12.64%, becomes 63.08% as SiO _2, was the molar ratio (Al / Si) = 0.24. On the other hand, the zeta potential was -33.84MV), calcium carbonate, calcium phosphate and hydroxyapatite The endotoxin adsorptive capacity was examined.

<Endotoxin adsorption test method>
Weigh accurately 5 mg of LPS (Lipopolysaccharide for biochemistry, manufactured by Wako Pure Chemical Industries, Ltd.), add distilled water for injection and dissolve to make 1000 mL. Weigh 200 mL of this solution with a graduated cylinder and add distilled water for injection to 1000 mL to obtain an endotoxin solution.

  Add 100 mL of endotoxin solution to a 200 mL beaker, add 0.1 g or 0.05 g of each powder sample with stirring, and stir for 10 minutes. After 10 minutes, filter with a 0.2 μm membrane filter and collect the filtrate. Dilute the filtrate exactly 500 times or 5000 times with distilled water for injection to make a sample solution.

  For each sample solution, operate according to the JP, General Test Method, Endotoxin Test Method, Kinetic-colorimetric method of optical measurement method (equipment: Well Reader SK601 (Seikagaku Corporation) wavelength: 405 nm), Quantify the endotoxin concentration in the sample solution. A blank test is performed in the same manner to determine the initial endotoxin concentration.

The test results are shown in Table 1 below.

(Test Example 2)
For the products of Examples 1 to 4, endotoxin adsorption ability was examined by changing the amount of sample added. The endotoxin adsorption test method was the same as in Test Example 1.

The test results are shown in Table 2 below.

(result)
As is apparent from the results of Test Examples 1 and 2, the endotoxin adsorption ability in Examples 1 to 4 of the present invention is much superior to other powders (adsorbents), and is extremely small as 0.05%. Adsorbed endotoxin at a high concentration (630,000EU / L) with a large amount of adsorbent, and could easily remove endotoxin. The endotoxin concentration of the injection is required to be 250 EU / L or less, and the endotoxin adsorption / removal agent of the present invention that can lower the endotoxin concentration to below the detection sensitivity sufficiently satisfies this requirement. .

Claims (2)

An endotoxin adsorption / removal agent characterized by containing aluminum silicate as an active ingredient and having an electrokinetic potential (zeta potential) when the aluminum silicate is dispersed in an aqueous solvent as a positive charge . The method for producing an endotoxin adsorption / removal agent according to claim 1, comprising a step of obtaining an aluminum silicate coprecipitate using an aluminum salt and colloidal silk as a silicon source.